Development and validation of pharmacokinetic assays for OBI-992, a TROP2-directed ADC: Application to human pharmacokinetic study

OBI-992 is a novel TROP2-targeted antibody–drug conjugate (ADC) composed of an anti-TROP2 monoclonal antibody conjugated to exatecan, a potent topoisomerase I (TOP1) inhibitor, via an enzyme-cleavable hydrophilic linker. OBI-992 is being assessed for its safety, tolerability, and pharmacokinetics in a Phase 1/2 clinical trial (NCT06480240). To characterize the pharmacokinetics of OBI-992, electrochemiluminescence (ECL) assays were developed and validated for the quantitative measurement of total antibody (TAb) and conjugated antibody (ADC) in human serum; an LC–MS/MS method was used to quantify the concentration of unconjugated exatecan in human plasma. To mitigate variability in ligand-binding assays (LBAs) due to different binding affinities of anti-payload antibodies to the reversible lactone–carboxylate forms of exatecan, a pH-optimized buffer system (pH 9.5) was implemented in the ADC assay. This approach enabled direct assessment of ADC stability and improved assay consistency without the need for acid pre-treatment. All methods were vigorously evaluated in compliance with FDA/ICH M10 guidelines, assessing accuracy, precision, dilution linearity, selectivity and stability. The methods demonstrated excellent sensitivity with validated quantitative ranges of 30–2500 ng/mL for TAb, 10–1000 ng/mL for ADC, and 10–10,000 pg/mL for unconjugated exatecan. All validation parameters met the predefined acceptance criteria, demonstrating the reliability and robustness of the methods. These validated methods were successfully applied to characterize the pharmacokinetic profile of OBI-992 in first-in-human study. Moreover, the pH-optimized ADC assay developed for OBI-992 may serve as a valuable precedent for future ADC programs facing similar analytical challenges with camptothecin (CPT)-based topoisomerase I inhibitor payloads.